Binding of the cyclic-AMP receptor protein (CRP) to specific DNA sequences stimulates transcription of most catabolic operons while repressing the activity of several others, including autogenous repression of its own structural crp gene. The positive regulatory aspect of CRP binding has been well investigated and the mechanism elucidated. However, the exact mechanism explaining how CRP binding inhibits transcription of its own structural gene remains unknown. To begin to understand a mechanism that can allow CRP binding to stimulate some operons while inhibiting its own, CRP binding mechanisms will be compared at a positively (lac) and a negatively (crp) effected promoter. To accomplish this objective, CRP interactions with the crp and lac promoters will be characterized in vitro using the polyacrylamide gel mobility shift assay (GMSA) to resolve the various CRP-DNA complexes that occur in solution. Specifically, this technique will allow determination of the stoichiometry, relative binding affinities and cooperativity. To determine how CRP binding effects the structure of the crp and lac promoters, CRP-promoter complexes will be examined using sedimentation velocity experiments. All of these experiments will analyze CRP-promoter complexes occurring at equilibrium. To examine the equilibrium aspects of in vivo CRP binding more closely, a computer assisted analysis of CRP binding to four different operons (lac, mal, gal, crp) has been performed. Specifically, the effects of [cAMP], [CRP] and [non-specific genomic DNA] on CRP binding have been examined.